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1.
BMC Infect Dis ; 21(1): 4, 2021 Jan 04.
Artículo en Inglés | MEDLINE | ID: mdl-33397312

RESUMEN

BACKGROUND: Tuberculous pleural effusion (TPE) is the most common extrapulmonary manifestation and may have lasting effect on lung function. However conventional diagnostic tests for TPE register multiple limitations. This study estimates diagnostic efficacy of the interferon gamma release assay (IGRA: T-SPOT.TB) in TPE patients of different characteristics. METHODS: We performed a prospective, single-centre study including all suspected pleural effusion patients consecutively enrolled from June 2015 to October 2018. Through receiver operating characteristic (ROC) curves, technical cut-offs and the utility of T-SPOT on pleural fluid (PF) were determined and analysed. Logistic regression analysis was performed to obtain the independent risk factors for TPE, and evaluated the performance of the T-SPOT assay stratified by risk factors in comparison to ADA. RESULTS: A total of 601 individuals were consecutively recruited. The maximum spot-forming cells (SFCs) of early secretory antigenic target-6 (ESAT-6) and culture filtrate protein-10 (CFP-10) in the PF T-SPOT assay had the best diagnostic efficiency in our study, which was equal to ADA (0.885 vs 0.887, P = 0.957) and superior to peripheral blood (PB), with a sensitivity of 83.0% and a specificity of 83.1% (The cut-off value was 466 SFCs/106 mononuclear cells). Among the TPE patients with low ADA (< 40 IU/L), the sensitivity and specificity of PF T-SPOT were still 87.9 and 90.5%, respectively. The utility of ADA was negatively related to increasing age, but the PF T-SPOT test had a steady performance at all ages. Age (< 45 yrs.; odds ratio (OR) = 5.61, 95% confidence interval (CI) 3.59-8.78; P < 0.001), gender (male; OR = 2.68, 95% CI 1.75-2.88; P < 0.001) and body mass index (BMI) (< 22; OR = 1.93, 95% CI 1.30-2.88; P = 0.001) were independently associated with the risk of TB by multivariate logistic regression analysis. Notably, when stratified by risk factor, the sensitivity of PF T-SPOT was superior to the sensitivity for ADA (76.5% vs. 23.5%, P = 0.016) and had noninferior specificity (84.4% vs. 96.9%, P = 0.370). CONCLUSIONS: In conclusion, the PF T-SPOT assay can effectively discriminate TPE patients whose ADA is lower than 40 IU/L and is superior to ADA in unconventional TPE patients (age ≥ 45 yrs., female or BMI ≥ 22). The PF T-SPOT assay is an excellent choice to supplement ADA to diagnose TPE.


Asunto(s)
Adenosina Desaminasa/análisis , Pruebas Diagnósticas de Rutina/métodos , Ensayos de Liberación de Interferón gamma/métodos , Mycobacterium tuberculosis/genética , Derrame Pleural/diagnóstico , Derrame Pleural/epidemiología , Tuberculosis Pleural/diagnóstico , Tuberculosis Pleural/epidemiología , Adenosina Desaminasa/sangre , Adulto , Anciano , Beijing/epidemiología , Exudados y Transudados/química , Exudados y Transudados/microbiología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Mycobacterium tuberculosis/aislamiento & purificación , Derrame Pleural/microbiología , Prevalencia , Estudios Prospectivos , Curva ROC , Factores de Riesgo , Sensibilidad y Especificidad , Esputo/química , Esputo/microbiología , Tuberculosis Pleural/microbiología
2.
J Int Med Res ; 49(1): 300060520984932, 2021 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-33461383

RESUMEN

OBJECTIVE: This study analyzed drug resistance and mutations profiles in Mycobacterium tuberculosis isolates in a surveillance site in Huairou District, Beijing, China. METHODS: The proportion method was used to assess drug resistance profiles for four first-line and seven second-line anti-tuberculosis (TB) drugs. Molecular line probe assays were used for the rapid detection of resistance to rifampicin (RIF) and isoniazid (INH). RESULTS: Among 235 strains of M. tuberculosis, 79 (33.6%) isolates were resistant to one or more drugs. The isolates included 18 monoresistant (7.7%), 19 polyresistant (8.1%), 28 RIF-resistant (11.9%), 24 multidrug-resistant (MDR) (10.2%), 7 pre-extensively drug-resistant (XDR, 3.0%), and 2 XDR strains (0.9%). A higher rate of MDR-TB was detected among previously treated patients than among patients with newly diagnosed TB (34.5% vs. 6.8%). The majority (62.5%) of RIF-resistant isolates exhibited a mutation at S531L in the DNA-dependent RNA polymerase gene. Meanwhile, 62.9% of INH-resistant isolates carried a mutation at S315T1 in the katG gene. CONCLUSION: Our results confirmed the high rate of drug-resistant TB, especially MDR-TB, in Huairou District, Beijing, China. Therefore, detailed drug testing is crucial in the evaluation of MDR-TB treatment.

3.
Infect Immun ; 89(3)2021 Feb 16.
Artículo en Inglés | MEDLINE | ID: mdl-33318140

RESUMEN

The lack of efficacious vaccines against Mycobacterium tuberculosis (MTB) infection is a limiting factor in the prevention and control of tuberculosis (TB), the leading cause of death from an infectious agent. Improvement or replacement of the BCG vaccine with one that reliably protects all age groups is urgent. Concerns exist that antigens currently being evaluated are too homogeneous. To identify new protective antigens, we screened 1,781 proteins from a high-throughput proteome-wide protein purification study for antigenic activity. Forty-nine antigens (34 previously unreported) induced antigen-specific gamma interferon (IFN-γ) release from peripheral blood mononuclear cells (PBMCs) derived from 4,452 TB and suspected TB patients and 167 healthy donors. Three (Rv1485, Rv1705c, and Rv1802) of the 20 antigens evaluated in a BALB/c mouse challenge model showed protective efficacy, reducing lung CFU counts by 66.2%, 75.8%, and 60%, respectively. Evaluation of IgG2a/IgG1 ratios and cytokine release indicated that Rv1485 and Rv1705c induce a protective Th1 immune response. Epitope analysis of PE/PPE protein Rv1705c, the strongest candidate, identified a dominant epitope in its extreme N-terminal domain accounting for 90% of its immune response. Systematic preclinical assessment of antigens Rv1485 and Rv1705c is warranted.

4.
J Virol ; 2020 Nov 11.
Artículo en Inglés | MEDLINE | ID: mdl-33177200

RESUMEN

Influenza A viruses cause severe respiratory illnesses in humans and animals. Overreaction of the innate immune response to influenza infection results in hypercytokinemia, which is responsible for mortality and morbidity. However, the mechanism by which influenza induces hypercytokinemia is not fully understood. In this study, we established a mouse-adapted H9N2 virus MA01 to evaluate the innate immune response to influenza in the lung. MA01 infection caused high levels of cytokine release, enhanced pulmonary injury in mice, and upregulated CD83 protein in dendritic cells and macrophages in the lung. Influenza neuraminidase unmasked CD83 protein and contributed to high cytokine levels. Furthermore, we provide evidence that CD83 is a sialylated glycoprotein. Neuraminidase treatment enhanced LPS-stimulated NF-κB activation in RAW264.7 cells. Anti-CD83 treatment alleviated influenza virus-induced lung injury in mice. Our study indicates that influenza neuraminidase modulates CD83 status and contributes to the "cytokine storm", which may suggest a new approach to curb this immune injury.IMPORTANCE The massive release of circulating mediators of inflammation is responsible for lung injury during influenza A virus infection. This phenomenon refers to the"cytokine storm". However, the mechanism by which influenza induces"cytokine storm" is not fully understood. In this study, we have shown that neuraminidase unmasked CD83 protein in the lung and contributed to high cytokine levels. Anti-CD83 treatment could diminish immune damage to lung tissue. The NA-CD83 axis may represent a target for an interrupt of influenza induced lung damage.

5.
BMC Infect Dis ; 20(1): 657, 2020 Sep 07.
Artículo en Inglés | MEDLINE | ID: mdl-32894079

RESUMEN

BACKGROUND: Tuberculosis is still a significant diagnostic and therapeutic challenge with high proportion of smear- and culture- negative incidences worldwide. The conventional diagnostic tests are time-consuming and have a low sensitivity. Digital PCR is a novel technology which can detect target sequences with relatively low abundance and obtain the absolute copy numbers of the targets. METHODS: We assessed the accuracy of dPCR in TB diagnosis using more than 250 specimens, and for the first time, we selected M.tuberculosis-specific IS1081 in addition to widely used IS6110 as the amplification targets for dPCR. The quantification of target DNA was calculated using QuantaSoft Version 1.7.4.0917 (BioRad), and SPSS version 13.0 software (SPSS Inc., Chicago, IL, USA) was used for statistical analyses. RESULTS: IS6110-dPCR was more sensitive than IS1081, with the sensitivity and specificity accounting for 40.6 and 93.4% respectively. When we classified the TB patients by personal factors for high copy number of M.tuberculosis derived DNA in plasma: bilateral TB, extrapulmonary TB and disseminated TB, the sensitivity of both IS6110- and IS1081- dPCR was the highest in patients with disseminated TB (IS6110, 100%; IS1081, 68.8%), while their sensitivity was a bit higher in patients with extrapulmonary TB (IS6110, 50.0%; IS1081, 39.3%) than that in bilateral TB (IS6110, 43.3%; IS1081, 33.3%). Compared with traditional TB diagnostic tests, joint detection IS6110 & IS1081-dPCR was not as sensitive as smear microscope or mycobacterial culture, but it was higher than IS6110 qPCR (p < 0.05) and was able to detect 47.4% of smear-negative TB patients. CONCLUSION: Our study suggested that plasma IS6110-dPCR is a rapid, moderate accurate and less-invasive method to detect M.tuberculosis DNA in plasma of TB patients and IS6110 & IS1081-dPCR has a potential to aid diagnosis of smear-negative TB.


Asunto(s)
Elementos Transponibles de ADN/genética , ADN Bacteriano/genética , Mycobacterium tuberculosis/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/métodos , Tuberculosis Pulmonar/diagnóstico , Adolescente , Adulto , Anciano , ADN Bacteriano/sangre , Exactitud de los Datos , Femenino , Humanos , Masculino , Persona de Mediana Edad , Sensibilidad y Especificidad , Adulto Joven
6.
iScience ; 23(7): 101288, 2020 Jul 24.
Artículo en Inglés | MEDLINE | ID: mdl-32622265

RESUMEN

IgE and IgG1 production in the type 2 immune response is the characteristic feature of an allergic reaction. However, whether bacterial molecules modulate IgE and IgG1 production remains obscure. Here, we demonstrate that the bacterial quorum sensing molecules acyl homoserine lactones (AHLs) induce IgE and IgG1 production by activating the RARE (retinoic acid response element) response in dendritic cells (DCs) in vivo. DC-specific knockout of the retinoic acid transcriptional factor Rara diminished the AHL-stimulated type 2 immune response in vitro. AHLs altered DC phenotype, upregulated OX40L and IFN-I signature, and promoted T helper 2 cell differentiation in vitro. Finally, AHLs activated the RARE response by inhibiting AKT phosphorylation in vitro, as the AKT agonists IGF-1 and PDGF abolished the effect of AHLs on the RARE response. This study demonstrates a mechanism by which AHLs drive allergic airway inflammation through activating retinoic acid signaling in DCs.

7.
Infect Genet Evol ; 84: 104363, 2020 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-32413573

RESUMEN

Mycobacterium tuberculosis (M. tuberculosis) Lineage 4 (L4) is frequently prevailing in Western regions of China, where the tuberculosis incidence rate is high. However, the epidemiology characteristics of M. tuberculosis L4 in China remain poorly understood. Here, the 15-loci Variable number of tandem repeats (VNTR) patterns of 975 L4 isolates from a National Survey of Tuberculosis in China were used to construct a Minimum Spanning Tree (MST), which divided the 975 isolates into 5 major clonal complexes (CC; named CC1 to CC5). We found that the CCs of M. tuberculosis L4 were nationally distributed, geographically restricted, and different in epidemiology characteristics. For example, CC1 was mainly concentrated in East and Central China and significantly related to the farmer occupation and income of an individual (>4200 yuan) (p < .05); CC5 was mainly distributed in Southwest China and was associated with ethnic minorities. Notably, using whole genome sequencing (WGS) data of 141 strains that matched our samples, we found that both CC1 and CC5 were mapped to the sublineage L4.5. Nevertheless, due to the difference of geographical distribution, the epidemiology characteristics of these CCs were largely different. We found that income and occupation significantly contributed to the odds of infection by CC1 to CC5. Consequently, our findings revealed the epidemiology characteristics of the CCs of M. tuberculosis L4, and will help in the formulation of more effective intervention measures in line with regional specifications and patient characteristics in China.

8.
Proteomics Clin Appl ; 14(1): e1900001, 2020 01.
Artículo en Inglés | MEDLINE | ID: mdl-31715074

RESUMEN

PURPOSE: To identify potential protein biomarkers for distinguishing tuberculosis plural effusion (TBPE) from malignant plural effusion (MPE). EXPERIMENTAL DESIGN: Five independent samples from each group (TBPE and MPE) are enrolled for label-free quantitative proteomics analyses. The differentially expressed proteins are validated by western blot and ELISA. Logistic regression analysis is used to obtain the optimal diagnostic model. RESULTS: In total, 14 proteins with significant difference are identified between TBPE and MPE. Seven differentially expressed proteins are validated using western blot, and the expression patterns of these seven proteins are similar with those in proteomics analysis. Statistically significant differences in four proteins (AGP1, ORM2, C9, and SERPING1) are noted between TBPE and MPE in the training set (n = 230). Logistic regression analysis shows the combination of AGP1-ORM2-C9 presents a sensitivity of 73.0% (92/126) and specificity of 89.4% (93/104) in discriminating TBPE from MPE. Additional validation is performed to evaluate the diagnostic model in an independent blind testing set (n = 80), and yielded a sensitivity of 74.4% (32/43) and specificity of 91.9% (34/37) in discriminating TBPE from MPE. CONCLUSION: The study uncovers the proteomic profiles of TBPE and MPE, and provides new potential diagnostic biomarkers for distinguishing TBPE from MPE.


Asunto(s)
Derrame Pleural Maligno/genética , Derrame Pleural/genética , Proteoma/genética , Tuberculosis/genética , Biomarcadores de Tumor/genética , Diagnóstico Diferencial , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Masculino , Persona de Mediana Edad , Proteínas de Neoplasias/genética , Derrame Pleural/diagnóstico , Derrame Pleural/patología , Derrame Pleural Maligno/diagnóstico , Derrame Pleural Maligno/patología , Proteómica , Tuberculosis/diagnóstico , Tuberculosis/patología
9.
Clin Lab ; 65(10)2019 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-31625364

RESUMEN

BACKGROUND: Interferon-gamma release assay (T-SPOT.TB) has the theoretical possibility of discriminating TB from most non-tuberculous mycobacteria (NTM) infections, but there are limited reports on the use of T-SPOT.TB for diseases due to NTM in high TB burden country. The aim of the present study was to assess the utility of T-SPOT.TB in patients with NTM pulmonary disease. METHODS: Clinical parameters and laboratory characteristics of patients with NTM pulmonary disease between July 2011 and Jan 2017 were investigated retrospectively and comprehensively reviewed. RESULTS: A total of 127 patients with NTM pulmonary disease were retrospectively reviewed. Seven NTM species were isolated from 115 patients, and the most common species were M. intracellulare (48.7%, 56/115) and M. abscessus (34.8%, 40/115). NTM isolates were mainly prevalent in people aged 50 years or older (73.0%). The overall positive rate of T-SPOT.TB test was 29.6% (24/81). In patients infected with NTM sharing the RD1 region of Mycobacterium tuberculosis (M. TB), 50% (3/6) were positive in the T-SPOT.TB test, whereas 28.0% (21/75) was positive in the group with NTM not sharing the RD1 region of M. TB. No significant difference was detected in the positive rate of T-SPOT.TB between definite (28.3%, 15/53) and probable disease (32.1%, 9/28). CONCLUSIONS: Our data indicated a relatively high positive rate of T-SPOT.TB test in patients infected with NTM not sharing the RD1 region of M. TB. Thus, T-SPOT.TB test displays a limited ability in differentiating TB infection from NTM disease in a high TB burden country.


Asunto(s)
Ensayos de Liberación de Interferón gamma/métodos , Infecciones por Mycobacterium no Tuberculosas/diagnóstico , Micobacterias no Tuberculosas/aislamiento & purificación , Tuberculosis Pulmonar/diagnóstico , Tuberculosis/diagnóstico , Adulto , Anciano , Anciano de 80 o más Años , Diagnóstico Diferencial , Femenino , Humanos , Masculino , Persona de Mediana Edad , Infecciones por Mycobacterium no Tuberculosas/sangre , Infecciones por Mycobacterium no Tuberculosas/microbiología , Micobacterias no Tuberculosas/clasificación , Micobacterias no Tuberculosas/fisiología , Reproducibilidad de los Resultados , Estudios Retrospectivos , Sensibilidad y Especificidad , Tuberculosis/sangre , Tuberculosis/microbiología , Tuberculosis Pulmonar/sangre , Tuberculosis Pulmonar/microbiología
10.
Dose Response ; 17(4): 1559325819882873, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31662712

RESUMEN

Backgrounds: This study compared analgesic effects and µ-opioid receptor expression levels during long-term intraperitoneal and intrathecal treatment in a bone cancer pain rat. Methods: Twenty-four female Sprague-Dawley rats were injected Walker 256 tumor cells into the femur to create a bone cancer pain model. The control group was injected with saline intraperitoneally and intrathecally. The intraperitoneal group was injected with morphine intraperitoneally and saline intrathecally. The intrathecal group was injected saline intraperitoneally and morphine intrathecally. Changes in pain threshold, µ-opioid receptor expression levels in spinal cord, and tumor tissue were compared between 3 groups. Results: The intrathecal morphine group and the intraperitoneal group showed no difference in analgesia effects (P > .05). Western blot and immunohistochemical staining of µ-opioid receptors demonstrated that its level in the intrathecal group was significantly lower than the intraperitoneal group (P < .05) and without significant difference with the control group (P > .05). The expression levels of µ-opioid receptor in the spinal cord tissue did not reveal a difference among these 3 groups (P > .05). Conclusion: Intrathecal group and intraperitoneal group showed significant difference in µ-opioid receptor expressions although with no difference in analgesia effects. Long-term intrathecal morphine administration provided similar analgesia compared to systemic morphine.

11.
Artículo en Inglés | MEDLINE | ID: mdl-31572691

RESUMEN

Tuberculous meningitis (TBM) is the most common and severe form of central nervous system tuberculosis. Due to the non-specific clinical presentation and lack of efficient diagnosis methods, it is difficult to discriminate TBM from other frequent types of meningitis, especially viral meningitis (VM). In order to identify the potential biomarkers for discriminating TBM and VM and to reveal the different pathophysiological processes between TBM and VM, a genome-wide miRNA screening of PBMCs from TBM, VM, and healthy controls (HCs) using microarray assay was performed (12 samples). Twenty-eight differentially expressed miRNAs were identified between TBM and VM, and 11 differentially expressed miRNAs were identified between TBM and HCs. The 6 overlapping miRNAs detected in both TBM vs. VM and TBM vs. HCs were verified by qPCR analysis and showed a 100% consistent expression patterns with that in microarray test. Statistically significant differences of 4 miRNAs (miR-126-3p, miR-130a-3p, miR-151a-3p, and miR-199a-5p) were further confirmed in TBM compared with VM and HCs in independent PBMCs sample set (n = 96, P < 0.01). Three of which were also showed significantly different between TBM and VM in CSF samples (n = 70, P < 0.05). The receiver operating characteristic curve (ROC) analysis showed that the area under the ROC curve (AUC) of these 4 miRNAs in PBMCs were more than 0.70 in discriminating TBM from VM. Combination of these 4 miRNAs could achieve better discriminative capacity [AUC = 0.893 (0.788-0.957)], with a sensitivity of 90.6% (75.0-98.0%), and a specificity of 86.7% (69.3-96.2%). Additional validation was performed to evaluate the diagnostic panel in another independent sample set (n = 49), which yielded a sensitivity of 81.8% (9/11), and specificity of 90.0% (9/10) in distinguishing TBM and VM, and a sensitivity of 81.8% (9/11), and a specificity of 84.6% (11/13) in discriminating TBM from other non-TBM patients. This study uncovered the miRNA profiles of TBM and VM patients, which can facilitate better understanding of the pathogenesis involved in these two diseases and identified 4 novel miRNAs in distinguishing TBM and VM.


Asunto(s)
Biomarcadores/sangre , Leucocitos Mononucleares/patología , Meningitis Viral/diagnóstico , MicroARNs/sangre , Tuberculosis Meníngea/diagnóstico , Adolescente , Adulto , Anciano , Diagnóstico Diferencial , Femenino , Perfilación de la Expresión Génica , Humanos , Masculino , Meningitis Viral/patología , Análisis por Micromatrices , Persona de Mediana Edad , Curva ROC , Reacción en Cadena en Tiempo Real de la Polimerasa , Sensibilidad y Especificidad , Tuberculosis Meníngea/patología , Adulto Joven
12.
Tuberculosis (Edinb) ; 118: 101861, 2019 09.
Artículo en Inglés | MEDLINE | ID: mdl-31526947

RESUMEN

Histone deacetylase inhibitors (HDACi), a novel class of anti-cancer drug, have been recently reported to suppress host immunity and increase susceptibility to infection. Tuberculosis, a leading infectious disease killer caused by Mycobacterium tuberculosis (M.tb), is basically the product of the interaction between bacterial virulence and host resistance. However, the effects of HDACi in host immunity against M.tb is largely unknown. In this study, we found that HDACi including Trichostatin A (TSA) and suberoylanilide hydroxamic acid (SAHA) significantly impaired phagocytosis and killing activity of macrophage. In line with these findings, we noted that M.tb induced reactive oxygen species (ROS) production and autophagy are significantly suppressed by TSA. Transcriptome analysis revealed that the suppression of autophagy by TSA might due to its inhibiting autophagy-regulating genes such as CACNA2D3, which regulates intracellular Ca2+ levels. Finally, we confirmed that HDACi including TSA and SAHA significantly exacerbated the histopathological damage and M.tb load in the lung of M.tb infected mice. Taken together, our results indicated that HDACi at least TSA and SAHA significantly impaired macrophage immunity against M.tb and therefore increase susceptibility to TB, our findings raised the concern that the potential side effects of HDACi on latent TB reactivation should be considered in clinic.


Asunto(s)
Inhibidores de Histona Desacetilasas/farmacología , Mycobacterium tuberculosis/efectos de los fármacos , Fagocitosis/efectos de los fármacos , Tuberculosis/inmunología , Animales , Autofagia/efectos de los fármacos , Autofagia/genética , Carga Bacteriana/efectos de los fármacos , Células Cultivadas , Recuento de Colonia Microbiana , Susceptibilidad a Enfermedades , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Regulación de la Expresión Génica/inmunología , Interacciones Microbiota-Huesped/efectos de los fármacos , Interacciones Microbiota-Huesped/genética , Interacciones Microbiota-Huesped/inmunología , Humanos , Ácidos Hidroxámicos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Mycobacterium tuberculosis/crecimiento & desarrollo , Mycobacterium tuberculosis/inmunología , Mycobacterium tuberculosis/aislamiento & purificación , Especies Reactivas de Oxígeno/metabolismo , Tuberculosis/microbiología , Vorinostat/farmacología
13.
Front Microbiol ; 10: 1174, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31191492

RESUMEN

Tuberculosis (TB) has been the leading lethal infectious disease worldwide since 2014, and about one third of the world's population has a latent TB infection (LTBI). This is largely attributed to the difficulties in diagnosis and treatment of TB and LTBI patients. Exosomes offer a new perspective on investigation of the process of TB infection. In this study, we performed small RNA sequencing to explore small RNA profiles of serum exosomes derived from LTBI and TB patients and healthy controls (HC). Our results revealed distinct miRNA profile of the exosomes in the three groups. We screened 250 differentially expressed miRNAs including 130 specifically expressed miRNAs. Some miRNAs were further validated to be specifically expressed in LTBI (hsa-let-7e-5p, hsa-let-7d-5p, hsa-miR-450a-5p, and hsa-miR-140-5p) and TB samples (hsa-miR-1246, hsa-miR-2110, hsa-miR-370-3P, hsa-miR-28-3p, and hsa-miR-193b-5p). Additionally, we demonstrated four expression panels in LTBI and TB groups, and six expression patterns among the three groups. These specifically expressed miRNAs and differentially expressed miRNAs in different panels and patterns provide potential biomarkers for detection/diagnosis of latent and active TB using exosomal miRNAs. Additionally, we also discovered plenty of small RNAs derived from genomic repetitive sequences, which might play roles in host immune responses along with Mtb infection progresses. Overall, our findings provide important reference and an improved understanding about miRNAs and repetitive region-derived small RNAs in exosomes during the Mtb infectious process, and facilitate the development of potential molecular targets for detection/diagnosis of latent and active tuberculosis.

14.
Infect Genet Evol ; 72: 183-190, 2019 08.
Artículo en Inglés | MEDLINE | ID: mdl-31242975

RESUMEN

Despite technical advances in introducing genomic deletions and modulating gene expression, direct inactivation of essential genes in mycobacteria remains difficult. In this study, we described clustered regularly interspaced short palindromic repeat interference (CRISPRi) technology to repress the expression of sepF (MSMEG_4219) based on nuclease-deficient CRISPR-associated protein 9 (Cas9) and small guide RNA (sgRNA) specific to the target sequence in Mycobacterium smegmatis. Using this CRISPRi approach, we achieved the repression of sepF by up to 98% in M. smegmatis without off-target effects. The depleted Msm_sepF strains resulted in growth and morphology changes including elongated, filamentous and branched bacterial cells, but the levels of the interacting partners ftsZ and murG were not modified in M. smegmatis. The sepF gene was proven to be an essential gene in M. smegmatis. This study provided an improved and detailed technical procedure for the application of CRISPRi technology in mycobacteria, and this approach was demonstrated to be a simple and efficient tool for regulating the expression of essential genes in M. smegmatis.


Asunto(s)
Sistemas CRISPR-Cas , Edición Génica , Técnicas de Silenciamiento del Gen , Mycobacterium smegmatis/genética , Proteínas Bacterianas/genética , Proteína 9 Asociada a CRISPR , Sistemas CRISPR-Cas/efectos de los fármacos , Sistemas CRISPR-Cas/genética , Edición Génica/métodos , Genes Esenciales , Mycobacterium smegmatis/efectos de los fármacos , Mycobacterium smegmatis/crecimiento & desarrollo
15.
Artículo en Inglés | MEDLINE | ID: mdl-30984625

RESUMEN

Tuberculosis (TB) is still a serious threat to human health which is caused by mycobacterium tuberculosis (Mtb). The main reason for failure to eliminate TB is lack of clearly understanding the molecular mechanism of Mtb pathogenesis. Determining human Mtb-interacting proteins enables us to characterize the mechanism and identify potential molecular targets for TB diagnosis and treatment. However, experimentally systematic Mtb interactors are not readily available. In this study, we performed an unbiased, comprehensive two-way proteome microarray based approach to systematically screen global human Mtb interactors and determine the binding partners of Mtb effectors. Our results, for the first time, screened 84 potential human Mtb interactors. Bioinformatic analysis further highlighted these protein candidates might engage in a wide range of cellular functions such as activation of DNA endogenous promoters, transcription of DNA/RNA and necrosis, as well as immune-related signaling pathways. Then, using Mtb proteome microarray followed His tagged pull-down assay and Co-IP, we identified one interacting partner (Rv0577) for the protein candidate NRF1 and three binding partners (Rv0577, Rv2117, Rv2423) for SMAD2, respectively. This study gives new insights into the profile of global Mtb interactors potentially involved in Mtb pathogenesis and demonstrates a powerful strategy in the discovery of Mtb effectors.


Asunto(s)
Interacciones Huésped-Patógeno , Mycobacterium tuberculosis/inmunología , Mycobacterium tuberculosis/patogenicidad , Proteoma/análisis , Tuberculosis/inmunología , Tuberculosis/patología , Biología Computacional , Humanos , Análisis por Micromatrices , Mycobacterium tuberculosis/química , Análisis por Matrices de Proteínas , Unión Proteica , Mapeo de Interacción de Proteínas , Tuberculosis/microbiología
16.
Zhongguo Fei Ai Za Zhi ; 22(4): 216-222, 2019 Apr 20.
Artículo en Chino | MEDLINE | ID: mdl-31014439

RESUMEN

BACKGROUND: MicroRNA is a kind of single-stranded non-coding RNA whose length is about 22 nucleotides and its abnormal expression is related to disease closely. This study is aiming to explore the relative expression of miR-34b-3p and miR-302a-5p in the plasma of non-small cell lung cancer (NSCLC) patients and its clinical value. METHODS: The levels of miR-34b-3p and miR-302a-5p in plasma were detected by real-time polymerase chain reaction (RT-PCR) in 86 patients with NSCLC, 64 patients with pulmonary tuberculosis (PTB) and 39 healthy subjects. Analyze their value in diagnosing NSCLC by contrasting and combining carcino-embryonic antigen (CEA), neuron-specific enolase (NSE), and cytokeratin 19 fragments 21-1 (CYFRA21-1). RESULTS: The levels of plasma miR-34b-3p and miR-302a-5p in NSCLC group were significantly higher than those in the PTB group and the healthy group (P<0.05). In patients with NSCLC, the levels of plasma miR-34b-3p was correlated with the diameter of tumor (P<0.01). When using one plasma marker to diagnose NSCLC, miR-302a-5p had the highest sensitivity (82.6%) and CEA had the highest specificity (81.6%). While combined two plasma markers, miR-34b-3p+miR-302a-5p had the highest sensitivity (80.2%) and miR-34b-3p+CEA had the highest specificity (81.4%). As detected multiple markers, miR-302a-5p+NSE+CYFRA21-1 had the highest sensitivity (81.4%) and miR-34b-3p+CEA+NSE had the highest specificity (90.3%). The combination of miR-34b-3p, miR-302a-5p and CEA obtained the highest area under the curve (AUC), which was 0.832. Logistic regression model indicated that miR-34b-3p was independent risk factor for NSCLC compared to control groups. CONCLUSIONS: Plasma miR-34b-3p and miR-302a-5p could be used as biological markers for the diagnosis of NSCLC.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/sangre , Carcinoma de Pulmón de Células no Pequeñas/diagnóstico , Neoplasias Pulmonares/sangre , Neoplasias Pulmonares/diagnóstico , MicroARNs/sangre , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Pronóstico
17.
Molecules ; 24(8)2019 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-30991677

RESUMEN

Four new compounds including two new sesquiterpenoid dimers, commiphoroids E (1) and F (2), a new triterpenoid (3), and a new sesquiterpenoid (4), along with three known terpenoids (5-7) were isolated from Resina Commiphora, whose structures were identified by NMR spectra, HRESIMS, and X-ray diffraction analysis. Compounds 1 and 2 both bear an O-bridge ring and feature a plausible [4 + 2] Diels-Alder cycloaddition reaction. Antimycobacterial activities show that all the tested compounds (200 µM) could inhibit the growth of both sensitive and clinically multi-drug resistant (MDR) isolated strains. In addition, cellular toxicity of the isolates against human cancer cells and THP-1 monocyte cells was examined.


Asunto(s)
Antituberculosos , Commiphora/química , Mycobacterium tuberculosis/crecimiento & desarrollo , Resinas de Plantas/química , Terpenos , Antituberculosos/química , Antituberculosos/farmacología , Humanos , Células THP-1 , Terpenos/química , Terpenos/farmacología
18.
Eur Respir J ; 52(6)2018 12.
Artículo en Inglés | MEDLINE | ID: mdl-30361241

RESUMEN

Latent tuberculosis infection (LTBI) management is now a critical component of the World Health Organization's End TB Strategy.In this randomised controlled trial (Chinese Clinical Trial Registry identifier ChiCTR-IOR-15007202), two short-course regimens with rifapentine plus isoniazid (a 3-month once-weekly regimen and a 2-month twice-weekly regimen) were initially designed to be evaluated for rural residents aged 50-69 years with LTBI in China.Due to the increasingly rapid growth and unexpected high frequency of adverse effects, the treatments were terminated early (after 8 weeks for the once-weekly regimen and after 6 weeks for the twice-weekly regimen). In the modified intention-to-treat analysis on the completed doses, the cumulative rate of active disease during 2 years of follow-up was 1.21% (14 out of 1155) in the untreated controls, 0.78% (10 out of 1284) in the group that received the 8-week once-weekly regimen and 0.46% (six out of 1299) in the group that received the 6-week twice-weekly regimen. The risk of active disease was decreased, with an adjusted hazard ratio of 0.63 (95% CI 0.27-1.43) and 0.41 (95% CI 0.15-1.09) for the treatments, respectively. No significant difference was found in the occurrence of hepatotoxicity (1.02% (13 out of 1279) versus 1.17% (15 out of 1279); p=0.704).The short regimens tested must be used with caution among the elderly because of the high rates of adverse effects. Further work is necessary to test the ultrashort regimens in younger people with LTBI.


Asunto(s)
Isoniazida/administración & dosificación , Tuberculosis Latente/tratamiento farmacológico , Rifampin/análogos & derivados , Anciano , Antibióticos Antituberculosos/uso terapéutico , China/epidemiología , Control de Enfermedades Transmisibles/métodos , Esquema de Medicación , Quimioterapia Combinada , Femenino , Humanos , Masculino , Persona de Mediana Edad , Mycobacterium tuberculosis/efectos de los fármacos , Modelos de Riesgos Proporcionales , Rifampin/administración & dosificación , Factores de Riesgo , Población Rural , Resultado del Tratamiento
19.
Mol Med Rep ; 18(2): 1551-1559, 2018 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-29901122

RESUMEN

Severe pulmonary tuberculosis (STB) is a life­threatening condition with high economic and social burden. The present study aimed to screen for distinct proteins in different stages of TB and identify biomarkers for a better understanding of TB progression and pathogenesis. Blood samples were obtained from 81 patients with STB, 80 with mild TB (MTB) and 50 healthy controls. Differentially expressed proteins were identified using liquid chromatography­tandem mass spectrometry­based label­free quantitative proteomic analysis. Functional and pathway enrichment analyses were performed for the identified proteins. The expression of potential biomarkers was further validated by western blot analysis and enzyme­linked immunosorbent assays. The accuracy, sensitivity and specificity for selected protein biomarkers in diagnosing STB were also evaluated. A total of 1,011 proteins were identified in all three groups, and 153 differentially expressed proteins were identified in patients with STB. These proteins were involved in 'cellular process', 'response to stimulus', 'apoptotic process', 'immune system process' and 'select metabolic process'. Significant differences in protein expression were detected in α­1­acid glycoprotein 2 (ORM2), interleukin­36α (IL­36α), S100 calcium binding protein A9 (S100­A9), superoxide dismutase (SOD)1 in the STB group, compared with the MTB and control groups. The combination of plasma ORM2, IL­36α, S100A9 and SOD1 levels achieved 90.00% sensitivity and 92.16% specificity to discriminate between patients with STB and MTB, and 89.66% sensitivity and 98.9% specificity to discriminate between patients with STB and healthy controls. ORM2, S100A9, IL­36α and SOD1 were associated with the development of TB, and have the potential to distinguish between different stages of TB. Differential protein expression during disease progression may improve the current understanding of STB pathogenesis.


Asunto(s)
Calgranulina B/genética , Interleucina-1/genética , Orosomucoide/genética , Superóxido Dismutasa-1/genética , Tuberculosis Pulmonar/diagnóstico , Tuberculosis Pulmonar/genética , Adulto , Anciano , Biomarcadores/sangre , Calgranulina B/sangre , Estudios de Casos y Controles , Progresión de la Enfermedad , Femenino , Perfilación de la Expresión Génica , Regulación de la Expresión Génica , Humanos , Interleucina-1/sangre , Masculino , Persona de Mediana Edad , Anotación de Secuencia Molecular , Orosomucoide/metabolismo , Mapeo de Interacción de Proteínas , Índice de Severidad de la Enfermedad , Superóxido Dismutasa-1/sangre , Tuberculosis Pulmonar/sangre , Tuberculosis Pulmonar/patología
20.
Front Microbiol ; 9: 1267, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-29951049

RESUMEN

The lack of effective differential diagnostic methods for active tuberculosis (TB) and latent infection (LTBI) is still an obstacle for TB control. Furthermore, the molecular mechanism behind the progression from LTBI to active TB has been not elucidated. Therefore, we performed label-free quantitative proteomics to identify plasma biomarkers for discriminating pulmonary TB (PTB) from LTBI. A total of 31 overlapping proteins with significant difference in expression level were identified in PTB patients (n = 15), compared with LTBI individuals (n = 15) and healthy controls (HCs, n = 15). Eight differentially expressed proteins were verified using western blot analysis, which was 100% consistent with the proteomics results. Statistically significant differences of six proteins were further validated in the PTB group compared with the LTBI and HC groups in the training set (n = 240), using ELISA. Classification and regression tree (CART) analysis was employed to determine the ideal protein combination for discriminating PTB from LTBI and HC. A diagnostic model consisting of alpha-1-antichymotrypsin (ACT), alpha-1-acid glycoprotein 1 (AGP1), and E-cadherin (CDH1) was established and presented a sensitivity of 81.2% (69/85) and a specificity of 95.2% (80/84) in discriminating PTB from LTBI, and a sensitivity of 81.2% (69/85) and a specificity of 90.1% (64/81) in discriminating PTB from HCs. Additional validation was performed by evaluating the diagnostic model in blind testing set (n = 113), which yielded a sensitivity of 75.0% (21/28) and specificity of 96.1% (25/26) in PTB vs. LTBI, 75.0% (21/28) and 92.3% (24/26) in PTB vs. HCs, and 75.0% (21/28) and 81.8% (27/33) in PTB vs. lung cancer (LC), respectively. This study obtained the plasma proteomic profiles of different M.TB infection statuses, which contribute to a better understanding of the pathogenesis involved in the transition from latent infection to TB activation and provide new potential diagnostic biomarkers for distinguishing PTB and LTBI.

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